Tumorigenicity of SV40 T antigen immortalized human prostate epithelial cells: Association with decreased epidermal growth factor receptor (EGFR) expression

Abstract

Our primary objectives were to: 1) develop a system for the study of prostatic tumor evolution; and 2) examine the role of the epidermal growth factor/epidermal growth factor receptor (EGF/EGFR) pathway in prostate tumor progression. Adult human prostate epithelial cells previously immortalized by transfection with the SV40 T antigen gene (P69SV40T) produced tumors in only 2/18 mice with a 6 month latency period. Reinjection of cells recovered from these tumors after 1 or 2 cycles of growth in nude mice produced tumors in 2/4 and 2/3 mice with markedly decreased latent intervals of 12, 25, 25 and 25 days each. The chromosomal complement of each tumor was human, consistently pseudodiploid, and retained the Y chromosome. In both anchorage-independent and adherent cell growth assays, EGF stimulated proliferation by approximately 2-fold in both the parental P69SV40T line and the tumor sublines. The tumor sublines expressed less EGFR protein than the parental line, as assessed by Western immunoblotting and flow cytometric analysis. Immunoprecipitation revealed increased production of the 18 and 25 kDa TGF-alpha precursors parallel to decreases in detectable EGFR. The growth of both the parental P69SV40T line and the tumor sublines was inhibited by a neutralizing antibody to TGF-alpha under serum-free defined conditions. Inclusion of the TGF-alpha neutralizing antibody consistently inhibited the proliferation of the tumor sublines more than P69SV40T in both proliferation and [3H]thymidine incorporation assays. This finding suggests that the increased tumorigenicity and decreased latent interval observed among the human prostate tumor cells is partially due to activation of the TGF-alpha/EGFR autocrine network.

Control of Protein Kinase C Activity, Phorbol Ester-induced Cytoskeletal Remodeling, and Cell Survival Signals by the Scaffolding Protein SSeCKS/GRAVIN/AKAP12

“…To strengthen this notion, we reproduced this analysis in human LNCaP cells, which exhibit severe down-regulation of SSeCKS (25-fold) compared with the untransformed, SV40 T antigen immortalized human epithelial cell line P69SV40T as we reported previously (48). Even with this down-regulation, SSeCKS is likely to exert some regulatory functions in this cell type because SSeCKS protein levels were still induced by culture confluence (Fig.…”

Unique aspects of mda-7/IL-24 antitumor bystander activity: establishing a role for secretion of MDA-7/IL-24 protein by normal cells

“…Human BxPC-3 pancreatic carcinoma cells, A549 human NSCLC and U87-MG malignant glioma cells were obtained from the ATCC and cultured in DMEM supplemented with 10% FBS. P69, an SV40-immortalized normal human prostate epithelial cell line, was provided by Dr Joy L Ware (Virginia Commonwealth University, Richmond, VA, USA) and grown under serum-free conditions as described previously (Bae et al, 1994). GBM6 is a primary malignant glioma cell line maintained in athymic nude mice (Yacoub et al, 2004).…”

Manganese superoxide dismutase overexpression inhibits the growth of androgen-independent prostate cancer cells

“…Even though these nonmalignant cells are not completely normal due to their immortalization, P69SV40T cells were considered as a good representation of normal prostate epithelial cells. The characterization of P69SV40T cell line was carried out by Bae et al (1994Bae et al ( , 1998. The MnSOD protein was measured with Western blot and the protein band corresponding to the MnSOD in PC-3 Wt cells was B50% of that of nonmalignant immortalized human prostate cells (P69SV40T) (Figure 1a).…”